METHODS: We conducted a two-stage time-stratified case-crossover study to examine the association between temperature and under-five mortality, spanning the period from 2014 to 2018 across all six regions in Malaysia. In the first stage, we estimated region-specific temperature-mortality associations using a conditional Poisson regression and distributed lag nonlinear models. We used a multivariate meta-regression model to pool the region-specific estimates and examine the potential role of local characteristics in the association, which includes geographical information, demographics, socioeconomic status, long-term temperature metrics, and healthcare access by region.
RESULTS: Temperature in Malaysia ranged from 22 °C to 31 °C, with a mean of 27.6 °C. No clear seasonality was observed in under-five mortality. We found no strong evidence of the association between temperature and under-five mortality, with an "M-" shaped exposure-response curve. The minimum mortality temperature (MMT) was identified at 27.1 °C. Among several local characteristics, only education level and hospital bed rates reduced the residual heterogeneity in the association. However, effect modification by these variables were not significant.
CONCLUSION: This study suggests a null association between temperature and under-five mortality in Malaysia, which has a tropical climate. The "M-" shaped pattern suggests that under-fives may be vulnerable to temperature changes, even with a small temperature change in reference to the MMT. However, the weak risks with a large uncertainty at extreme temperatures remained inconclusive. Potential roles of education level and hospital bed rate were statistically inconclusive.
METHODOLOGY/PRINCIPAL FINDINGS: Two ORFeome phage display libraries of the entire Leptospira spp. genomes from five local strains isolated in Malaysia and seven WHO reference strains were constructed. Subsequently, 18 unique Leptospira peptides were identified in a screen using a pool of sera from patients with acute leptospirosis. Five of these were validated by titration ELISA using different pools of patient or control sera. The diagnostic performance of these five peptides was then assessed against 16 individual sera from patients with acute leptospirosis and 16 healthy donors and was compared to that of two recombinant reference proteins from L. interrogans. This analysis revealed two peptides (SIR16-D1 and SIR16-H1) from the local isolates with good accuracy for the detection of acute leptospirosis (area under the ROC curve: 0.86 and 0.78, respectively; sensitivity: 0.88 and 0.94; specificity: 0.81 and 0.69), which was close to that of the reference proteins LipL32 and Loa22 (area under the ROC curve: 0.91 and 0.80; sensitivity: 0.94 and 0.81; specificity: 0.75 and 0.75).
CONCLUSIONS/SIGNIFICANCE: This analysis lends further support for using ORFeome phage display to identify pathogen-associated immunogenic peptides, and it suggests that this technique holds promise for the development of peptide-based diagnostics for leptospirosis and, possibly, of vaccines against this pathogen.